TY - JOUR
T1 - MHC class I stability is modulated by cell surface sialylation in human dendritic cells
AU - Silva, Zélia
AU - Ferro, Tiago
AU - Almeida, Danielle
AU - Soares, Helena
AU - Ferreira, José Alexandre
AU - Deschepper, Fanny M.
AU - Hensbergen, Paul J.
AU - Pirro, Martina
AU - van Vliet, Sandra J.
AU - Springer, Sebastian
AU - Videira, Paula A.
PY - 2020/3
Y1 - 2020/3
N2 - Maturation of human Dendritic Cells (DCs) is characterized by increased expression of antigen presentation molecules, and overall decreased levels of sialic acid at cell surface. Here, we aimed to identify sialylated proteins at DC surface and comprehend their role and modulation. Mass spectrometry analysis of DC’s proteins, pulled down by a sialic acid binding lectin, identified molecules of the major human histocompatibility complex class I (MHC-I), known as human leucocyte antigen (HLA). After desialylation, DCs showed significantly higher reactivity with antibodies specific for properly folded MHC-I-β2-microglobulin complex and for β2-microglobulin but showed significant lower reactivity with an antibody specific for free MHC-I heavy chain. Similar results for antibody reactivities were observed for TAP2-deficient lymphoblastoid T2 cells, which express HLA-A*02:01. Using fluorescent peptide specifically fitting the groove of HLA-A*02:01, instead of antibody staining, also showed higher peptide binding on desialylated cells, confirming higher surface expression of MHC-I complex. A decay assay showed that desialylation doubled the half-life of MHC-I molecules at cell surface in both DCs and T2 cells. The biological impact of DC´s desialylation was evaluated in co-cultures with autologous T cells, showing higher number and earlier immunological synapses, and consequent significantly increased production of IFN-γ by T cells. In summary, sialic acid content modulates the expression and stability of complex MHC-I, which may account for the improved DC-T synapses.
AB - Maturation of human Dendritic Cells (DCs) is characterized by increased expression of antigen presentation molecules, and overall decreased levels of sialic acid at cell surface. Here, we aimed to identify sialylated proteins at DC surface and comprehend their role and modulation. Mass spectrometry analysis of DC’s proteins, pulled down by a sialic acid binding lectin, identified molecules of the major human histocompatibility complex class I (MHC-I), known as human leucocyte antigen (HLA). After desialylation, DCs showed significantly higher reactivity with antibodies specific for properly folded MHC-I-β2-microglobulin complex and for β2-microglobulin but showed significant lower reactivity with an antibody specific for free MHC-I heavy chain. Similar results for antibody reactivities were observed for TAP2-deficient lymphoblastoid T2 cells, which express HLA-A*02:01. Using fluorescent peptide specifically fitting the groove of HLA-A*02:01, instead of antibody staining, also showed higher peptide binding on desialylated cells, confirming higher surface expression of MHC-I complex. A decay assay showed that desialylation doubled the half-life of MHC-I molecules at cell surface in both DCs and T2 cells. The biological impact of DC´s desialylation was evaluated in co-cultures with autologous T cells, showing higher number and earlier immunological synapses, and consequent significantly increased production of IFN-γ by T cells. In summary, sialic acid content modulates the expression and stability of complex MHC-I, which may account for the improved DC-T synapses.
KW - Antigen-presentation
KW - Cancer-vaccines
KW - Dendritic-cells
KW - Immunogenicity
KW - MHC-I
KW - T-cell response
UR - http://www.scopus.com/inward/record.url?scp=85081607435&partnerID=8YFLogxK
U2 - https://doi.org/10.3390/pharmaceutics12030249
DO - https://doi.org/10.3390/pharmaceutics12030249
M3 - Article
C2 - 32164343
SN - 1999-4923
VL - 12
JO - Pharmaceutics
JF - Pharmaceutics
IS - 3
M1 - 249
ER -